Functional genomics of the Clostridium thermocellum cellulosome

• Antragsteller: Dr. Wolfgang H. Schwarz
• Fachliche Zuordnung: Parasitologie und Biologie der Erreger tropischer Infektionskrankheiten
• Förderung: Förderung von 2002 bis 2007
• Projektkennung: Deutsche Forschungsgemeinschaft (DFG) - Projektnummer 5374166

Zusammenfassung der Projektergebnisse

In the course of the DFG project "Functional genomics of the Clostridium thermocellum cellulosome" the composition of the cellulosomes from the thermophilic anaerobic bacterium C. thermocellum was investigated. Cultures were grown on 4 different substrates and cellulosomes were prepared. Altogether 32 proteins were identified to be expressed and incorporated into the complex. The 19 components present in more then 3 (of 5) preparations were investigated in particular. The hitherto unknown proteins were expressed in E. coli. Extraordinary is Cel9R which is a processive endo-glucanase producing cellotetraose. Other enzymes with similar traits were also detected. All essential cellulase components are now available for a rational in vitro reconstitution of the cellulosome for an effective hydrolysis of crystalline cellulose. Analysis of the genome sequence of C. thermocellum resulted in a minimum of about 71 genes involved in cellulosome formation. Their sequences were analyzed for modular structure and potential function. Interesting components were biochemically characterized, e.g. a pectinase system. The list contains further potentially interesting enzyme components which should be analyzed in the future. The identification of the components essential for crystalline cellulose formation yielded in the scaffoldin CipA defective mutants which were used as a tool for the analysis of cellulosomal function. These mutants showed unequivocally the great effect of complex formation for the synergism of different cellulases and open a window for the development of dramatically more active cellulases for industrial purposes. A new active IS element (IS1447) was detected to be inserted in the mutant cipA genes.

Projektbezogene Publikationen (Auswahl)

• Schwarz, W.H. (2007) Bacterial cellulose hydrolysis in anaerobic environmental subsystems: Clostridium thermocellum - the cellulose cruncher. Eingeladener Plenarvortrag auf der Tagung "Incredible Anaerobes", Athens GA, USA, 02.-03.03.07
  
  
• Berger, E., Zhang, D., Zverlov, V.V. & Schwarz, W.H. (2007). Two noncellulosomal cellulases of Clostridium thermocellum, Cel9I and Cel48Y, hydrolyse crystalline cellulose synergistically. FEMS Microbiol. Let. 268: 194-201. (Siehe online unter: doi: 10.1111/j.1574- 6968.2006.00583.x)
  
  
• Schwarz, W. H. (2005) "Clostridium thermocellum cellulosome using genomic / proteomic approaches". Gordon Research Conference, Andover, NH, USA, Aug. 7.-12., 2005.
  
  
• Schwarz, W. H., Zverlov, V. V. (2005) Bacteria for the utilization of lignocellulosic biomass for fuel production. MEM-05 Symposium, München
  
  
• Schwarz, W.H. (2005) Microbiology of polysaccharide degradation and solvent production. Vortrag TU-Wien, Nov. 05
  
  
• Schwarz, W.H. (2006) Bakterieller Cellulose-Abbau: Lösungen für ein Problem? Vortrag Uni Stuttgart am 03.05.06.
  
  
• Schwarz, W.H. (2006). Präsentation "Cellulasen für spezielle Aufgaben" auf dem Arbeitskreis Bioraffinerie, DECHEMA, Frankfurt, 14.02.06.
  
  
• Schwarz, W.H., Zverlov, V.V. (2006). Protease inhibitors in bacteria: an emerging concept for the regulation of bacterial protein complexes? Molec. Microbiol. 60: 1323-1326. (Siehe online unter: doi: 10.1111/j.1365-2958.2006.05181.x)
  
  
• Schwarz, W.H., Zverlov, V.V., & Bahl, H. (2004). Extracellular glycosyl hydrolases from clostridia. Adv. Appl. Microbiol. 56, 215-261. ISSN 0065-2164 (Print)
  
  
• Zverlov V. V. , W. H. Schwarz (2005) Bacterial hydrolysis of lignocellulosic biomass: the Clostridium thermocellum cellulosome using genomic / proteomic approaches. Vortrag am IMG (Institute for Molecular Genetics, Academy of Sciences, Moscow). 05.09.05
  
  
• Zverlov, V. V., M. Hösl, E. Eckers, A. Pegel, W. H. Schwarz (2006) The major components in the Clostridium thermocellum cellulosome: proteins in four cellulosome preparations from different substrates. Kongress Clostridium IX, 19.05.05, Houston USA.
  
  
• Zverlov, V. V., Schwarz, W. H. (2005) New developments towards the understanding of the Clostridium thermocellum cellulosome. RITE International Symposium, International Workshop on Biorefinery, Kyoto, 9.-11.02.2005.
  
  
• Zverlov, V. V., Schwarz, W. H. (2005) The Clostridium thermocellum cellulosome: detection of the major components and characterization of three new hydrolytic enzymes. Jahrestagung der VAM und DGHM in Göttingen, 24.-27.09.2005.
  
  
• Zverlov, V. V., Velikodvorskaya, G. A., Schwarz, W. H. (2003). Two new cellulosome components encoded downstream of celI in the genome of Clostridium thermocellum: the non-processive endoglucanase CelN and the possibly structural protein CseP. Microbiol. 149: 515-524.
  
  
• Zverlov, V. V., W. H. Schwarz (2005) The Clostridium thermocellum cellulosome: detection of the major components and characterization of three new hydrolytic enzymes. MEM-05 Symposium, München
  
  
• Zverlov, V.V., & Schwarz, W.H. (2006). The C. thermocellum cellulosome: novel components and insights from the genomic sequence. In Cellulosome, chapt. VI, ed. Vladimir Uversky & Irina Kataeva. (In: Molecular Anatomy and Physiology of Proteins, edited by Vladimir Uversky). Nova Publishers Kataeva, I., & Ljungdahl, L. (eds.). ISBN 1-59454-950-8
  
  
• Zverlov, V.V., Kellermann, J., & Schwarz, W.H. (2005). Functional subgenomics of Clostridium thermocellum cellulosomal genes: Identification of the major catalytic components in the extracellular complex and detection of three new enzymes. Proteomics, 5. 3646-3653. Copyright © 2005 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim, DOI 10.1002/pmic.200401199. ISSN: 1615-9853 (print), 1615-9861 (online)
  
  
• Zverlov, V.V., Schantz, N., & Schwarz, W.H. (2005). A major new component in the cellulosome of Clostridium thermocellum is a processive endo-ß-1,4-glucanase producing cellotetraose. FEMS Microbiol. Let. 249: 353-358. Copyright © 2005 Federation of European Microbiological Societies Published by Elsevier B.V.. ISSN: 0378-1097 (Siehe online unter: doi: 10.1016/j.femsle.2005.06.037)
  
  
• Zverlov, V.V., Schmitt-Kopplin, P., Schantz, N., & Schwarz, W.H. (2005). Two new major subunits in the cellulosome of Clostridium thermocellum: xyloglucan hydrolase Xgh74A and endo-xylanase Xyn10D. Microbiol. 151: 3395-3401. ISSN 1350-0872. Copyright Society for General Microbiology, Reading, UK.
  
  
• Zverlov, V.V., Velikodvorskaya, G.A., and Schwarz, W. H. (2002). A newly described cellulosomal cellobiohydrolase, CelO, from Clostridium thermocellum: investigation of the exo-mode of hydrolysis, and binding capacity to crystalline cellulose. Microbiology 148:247-255.
  
  
• Zverlov, V.V., Velikodvorskaya, G.A., and Schwarz, W. H. (2002). A newly described cellulosomal cellobiohydrolase, CelO, from Clostridium thermocellum: investigation of the exo-mode of hydrolysis, and binding capacity to crystalline cellulose. Microbiology 148:247-255.
  
  
• Zverlov, V.V.1; Schwarz, W.H. (2005) The Clostridium thermocellum cellulosome: detection of the major components and characterization of three new hydrolytic enzymes. VAAM-Tagung, Göttingen 25.-27.8.05